Inhibition of cardiac myocyte apoptosis during ischemia-reperfusion and heart failure decreases myocardial damage, left ventricular remodeling, contractile dysfunction and, in some cases, mortality. Little is known, however, about the molecular regulation of apoptosis specifically in cardiac myocytes. The central death machinery has been highly conserved from worm to human and differs little among various cell types. Despite this, apoptosis is often regulated in a cell type- and stimulus-specific manner, the basis of which is poorly understood. ARC (Apoptosis Represser with a CARD (caspase recruitment domain)) is an endogenous inhibitor of apoptosis that is expressed primarily in cardiac and skeletal muscle. Our preliminary studies demonstrate that ARC suppresses both the death receptor and mitochondrial pathways through novel protein-protein interactions. In the death receptor pathway, ARC binds directly to Fas and FADD to inhibit the formation of the Death Inducing Signaling Complex which is required for death receptor signaling. In the mitochondrial pathway, ARC binds directly to Bax to inhibit Bax activation and translocation to the mitochondria in response to apoptotic stimuli. The ability of ARC to simultaneously antagonize both central death pathways is unique and suggests that this inhibitor may function as a master-repressor of apoptosis in cardiac myocytes. This application tests the central hypothesis that ARC serves as a critical regulator of cardiac myocyte apoptosis through Bax, Bak, and mitochondrial-dependent mechanisms. The objectives are to determine (a) the mechanisms by which ARC inhibits apoptotic pre-mitochondrial and mitochondrial events; and (b) the biological significance of this inhibition for cardiac myocyte survival in pathological contexts. Aim 1 seeks to delineate mechanisms by which ARC inhibits Bax translocation, Bak activation, and apoptotic mitochondrial events. Aim 2 uses Bax and Bak knockout mice to define the role of these downstream ARC effectors in acute ischemia-reperfusion injury and post-infarct remodeling in vivo. Aim 3 uses ARC knockout mice to determine if the absence of ARC causes a baseline cardiomyopathy and/or exacerbates acute ischemia reperfusion injury and post-infarct remodeling in vivo. In addition, this aim tests whether these phenotypes can be rescued by simultaneous deletion of Bax or Bak. These aims constitute a highly focused and integrated program that examines critical aspects of ARC from the single molecule to its role in the intact animal. The resulting information will advance our understanding of apoptosis in the myocardium and may provide the basis for novel cardiac-specific therapies for myocardial infarction and heart failure.